A collection of Hydnotrya confusa Spooner 1992 from Southwest Germany

Martin BEMMANN Ditte BANDINI Kleingemünderstraße 111 Panoramastr. 47 D-69118 Heidelberg D-69257 Wiesenbach [email protected] [email protected]

Ascomycete.org, 3 (3) : 55-60. Summary: A recent find of a hypogeous ascomycete from Germany that could be iden- Octobre 2011 tified as Hydnotrya confusa is reported and discussed.This species is described here for the first time from fresh material.

Keywords: Ascomycota, Pezizales, Discinaceae, hypogeous fungi.

Zusammenfassung: Ein aktueller Fund eines hypogäischen Ascomyzeten aus Süd- westdeutschland konnte als Hydnotrya confusa bestimmt werden. Die Art wird hier zum ersten Mal basierend auf Frischmaterial vorgestellt.

Schlüsselwörter: Ascomycota, Pezizales, Discinaceae, hypogäische Pilze.

Introduction the staining agents cotton blue (in lactic acid) and brilliant cresyl blue (aqueous) were used. Microphotography was Hydnotrya confusa was first described with a Latin diagno- done using a Nikon Coolpix 5000 through a Zeiss achromate 10x/0.22 and a Zeiss planapochromate 40x/1.0 Oel m.I. im- sis by SPOONER (1992). A detailed description with illustra- mersion objective. tions followed in PEGLER et al. (1993: 79, 82-83) using six collections from the UK (then herbarised as H. ploettneriana = H. michaelis) and separating this taxon from H. michaelis Habitat and H. cubispora. The Dutch mycologist G. A. de Vries sub- sequently examined his herbarium material and identified D-BW Schönbrunn, „Todtenbronnen“ (49°24’26” N, four collections from the Netherlands and one from Belgium 8°57’09” E, 361 m, MTB 6519/4/34), on boggy soil, partly as the same species previously classified as H. michaelis as churned up by wild boar, found on the surface in secondary well (DE VRIES, 2002). A provisional key to the European spe- position, 15.VII.2011. leg. D. Bandini, det. M. Bemmann, cies of Hydnotrya was provided by REIL (1999) and STIELOW et conf. B. Spooner (based on macro- and microphotographs). al. (2010), the latter key excluding H. confusa as being a non When the site was revisited by the authors on 31.VII.2011, Central European species then. 25 freshly grown semi-epigeous fruitbodies were observed in The genus Hydnotrya Berk. & Broome belonged, at the time situ. of Spooner’s publication, to the Helvellaceae. In the mean- The “Todtenbronnen” is a fen crossed by a small brook. The time, it was shifted to the Discinaceae by molecular means major, southern part of it is mostly an open area while the northern tail (where the site is situated) is covered by forest. (HANSEN & PFISTER, 2006; TEDERSOO et al., 2006; LÆSSØE & HAN- SEN, 2007). The new record of this species for Germany indicates that its distribution may be wider than known until now, and there may still exist some mislabelled specimens in different her- baria, as it was the case in the UK and the Netherlands.

Materials and methods

The first fruitbody was found on 15.VII.2011 to which most of the microscopical analysis refers. On 31.VII.2011, the site was revisited and another 25 fruitbodies could be observed. Their macro- and microscopical appearance (three fruitbo- dies tested) is included in the description below. The specimens were examined from fresh material, matu- red in a moist chamber for a period of up to two weeks at room temperature. Microscopy and all micro-measurements were carried out in tap water. For additional observations, Fig. 1 — The site. Photo: M. Bemmann. 55 Fig. 2 — Hydnotrya confusa. Photo: M. Bemmann. Fig. 3 — Hydnotrya confusa. Photo: D. Bandini. Ascomata in situ close to roots of Picea abies. Ascoma in situ.

Fig. 4 — Hydnotrya confusa. Photo: M. Bemmann. Fig. 5 — Hydnotrya confusa. Photo: M. Bemmann. Ascoma in situ with mycorrhizal roots (red arrow). Mycorrhizal roots.

On the site both banks of the brook are very boggy. Here the some Fraxinus excelsior. The place is very damp and surely dominating trees are Alnus, Fagus, Fraxinus, Betula and frequently flooded after heavy rainfall or snowmelt by the ad- Frangula. Further vegetation observed is Dryopteris, Pteri- jacent brook. Picea is most likely the mycorrhizal partner, as dium, Lycopus, Impatiens, Circaea, Scrophularia and diffe- the fruitbodies were found very close to their roots growing rent mosses, among them patches of Sphagnum. A few in the needle litter (figs. 2-3). Some ectomycorrhizal roots meters away from the brook the soil level is rising with a dif- could be observed next to the ascomata (figs. 4-5). ferent vegetation. A dense forest of Picea abies stretches on the left side with predominantly humid mossy soil. On the Description right side are growing Picea and Fagus with Vaccinium and mosses below. But also in the boggy part close to the brook some Picea are growing. The ascomata are of irregular shape ranging in size from ca. While the first specimen was found excavated by wild boars, 2 × 2 × 1.5 cm up to 4 × 4 × 2 cm. They are hollow, single- the other ascomata were found in situ in semi-epigeous po- chambered but cerebriformously folded, with a primary api- sition, erumpent from the soil and needle litter, and easily cal opening (fig. 6) and sometimes some smaller secondary visible due to their light colour but obviously ignored by the openings that could be traces of feeding by small rodents or boars. As the site was already revisited on 22.VII.2011 wi- snails. Such “secondary” openings were observed by Soeh- thout success, it is clear that these fructifications must have ner in his collection of Hydnotrya carnea as well (SOEHNER, grown within a week’s time. Some specimens were collec- 1951: 12 et seq.). Smaller, younger specimens sometimes ted for examination and herbarization while the rest was appear to have no opening at all. The smell is insignificant. kept in situ to ensure reproduction. However, one week later The colour of the outer surface is whitish to cream when only traces of some ascomata were left. young and gets yellowish with some few brown tints at age. The site where fresh ascomata were found covers an area of The texture of the surface is finely warty to tomentose. The c. 5 × 5 m (fig. 1). It is surrounded by young Picea abies and thickness of the flesh measures c. 1 mm. 56 Fig. 6 — Hydnotrya confusa. Photo: M. Bemmann. Fig. 7 — Hydnotrya confusa. Photo: M. Bemmann. Ascoma showing apical opening (red arrow). Ascoma in section.

The peridium in section (figs. 7-8) is yellowish with a lighter outer zone caused by the hyaline tomentum hyphae that are of cylindric to pyriform shape (fig. 16C). It is followed by a whitish subhymenium of variable thickness consisting of hyaline cells of irregular shape (textura angularis). The hy- menium is two-layered in colour, the one up to the ascus tips is yellowish in immature ascomata but deep orange in ma- ture ones due to the colour of the developed spores. The up- permost zone, composed of the heads of the hyaline paraphyses, appears again pure white when young but dull yellowish with age (figs. 8-9). Mature asci are cylindric, 270–360 × 31–36 µm, with uni- seriately arranged spores (figs. 9 and 12), immature asci are more clavate in shape, with irregular or biseriately arranged spores (those not featuring an exosporium yet) (figs. 10 and 16A). Immature ascospores are globose, hyaline, and are featu- ring regularly one oil-drop, that increases in size during ma- turation by nearly filling the whole spore lumen. Mature spores develop a lobate exosporium, golden brown with age and with a radially striate structure in optical section (figs. 13, 14 and 16D). The size of the spores is 33–47 × 25– 33 µm (incl. ornament) and 19.5–23.5 × 18.5–20.5 (excl. ornament). Most of the spores develop a more or less cir- cularly shaped exosporium but some appear rectangular in optical section resembling those of H. cubispora, maybe due to the limitation of horizontal space for growing within the ascus (fig. 11). The same observation was made by DE VRIES (2002: 203, fig. 1). However, spores featuring a rectangular exosporium were not recorded outside the asci. Paraphyses are hyaline, straight (5–6.5 µm in diam.), api- Fig. 8 — Hydnotrya confusa. Photo: M. Bemmann. cally slightly inflated (up to 8–10 (11) µm wide) and septate Section of an immature (top) and mature (bottom) ascoma. (figs. 15 and 16B). They are protruding from the asci by c. 150 µm. the present collection in few cases only, but sometimes, as DE VRIES (2002: 204) had already observed, in a tilted posi- Discussion tion (fig. 13).The width of the paraphyses in the present col- lection is even larger than in the examples described from Hydnotrya confusa differs from H. michaelis and H. cubis- the UK. pora mainly in the measurements of the asci and the para- The genus Hydnotrya surely deserves more research on physes and in the morphology of the spores. However, the taxonomical and molecular level with such a few collections observation of PEGLER et al. (2003: 82) that ascospores are known. As for H. confusa the authors would like to stress the placed “almost horizontal” within the asci could be noted in consideration given in PEGLER et al. (2003: 82 et seq.) that 57 Fig. 9 — Hydnotrya confusa. Photo: M. Bemmann. Section showing mature asci.

Fig. 10 — Hydnotrya confusa. Photo: M. Bemmann. Immature asci with globose spores.

Fig. 11 — Hydnotrya confusa. Photo: M. Bemmann. Fig. 12 — Hydnotrya confusa. Photo: M. Bemmann. Submature asci and spores. Hymenium with mature asci and spores. 58 Fig. 13 — Hydnotrya confusa. Photos: M. Bemmann. Mature spores inside asci in water (left) and cresyl blue (right). Fig. 15 — Hydnotrya confusa. Photo: M. Bemmann. Paraphyses.

Fig. 14 — Hydnotrya confusa. Photos: M. Bemmann. Mature spores. 59 B

A

Fig. 17 — “Hydnotrya carnea” after TICHOMIROW (1908a, 1908b)

C

D References

HANSEN K. & PFISTER D.H. 2006. — Systematics of the Pezizomy- cetes - the operculate discomycetes. Mycologia, 98 (6): 1029- 1040. LÆSSØE T. & HANSEN K. 2007. — Truffle trouble: what happened to the Tuberales? Mycological Research, 111 (9): 1075-1099. Fig. 16 — Hydnotrya confusa. Microscopic characters. PEGLER D.N., SPOONER B.M. & YOUNG T.W.K. 1993. — British Truf- Drawings: D. Bandini. fles. A revision of British hypogeous fungi. Kew. A. Immature and submature asci. B. Paraphyses. C. Tomentum hy- REIL P. 1999. — Hydnotrya cerebriformis, ein Fund im Schwarz- phae of the peridium. D. Mature spores wald. Deutsche Gesellschaft für Mykologie (ed.), Bemerkens- werte Pilze - Icones miscellaneae fungorum. Zeitschrift für Mykologie, Beiheft, 9: 113-118. doubtful taxa like H. soehneri could be conspecific. TICHOMI- SOEHNER E. 1951. — Tuberaceen-Studien III. Zeitschrift für Pilz- ROW (1908a, 588 et seq., Abb. 6; 1908b, 194 et seq., pl. V, kunde, 8: 7-16. 6) published some pictures of asci, paraphyses and spores SPOONER B. 1992. — A new species of Hydnotrya (Helvellaceae) from the British Isles. Kew Bulletin, 47 (3): 503. of a fungus he called "Hydnotrya carnea Zobel" from Rus- STIELOW B., BUBNER B., HENSEL G., MÜNZENBERGER B., HOFFMANN P., KLENK sian finds simultaneously in German and French pharma- H.P. & GÖKER M. 2010. — The neglected hypogeous fungus Hyd- cological journals, but did not give a description of the notrya bailii Soehner (1959) is a widespread sister taxon of Hyd- morphology of the ascocarps. The shape of the spores in his notrya tulasnei (Berk.) Berk. & Broome (1846). Mycological micro-drawings (fig. 17) resemble those of H. confusa. The Progress, 9 (2): 195-203. paraphyses are septate but appear to be shorter than the TEDERSOO L., HANSEN K., PERRY B.A. & KJØLLER R. 2006. — Molecular asci. This is maybe owed to his excessive treatment with che- and morphological diversity of pezizalean ectomycorrhiza. New micals as phenylhydracine, iodine and boiling in Fehling’s Phytologist, 170: 581-596. solution, as Tichomirow was not particularly interested in this TICHOMIROW A.W. 1908a. — Das Glykogen der Ascomycetenpilze in seinen Beziehungen zur Trehalose. Archiv der Pharmazie, 246 fungus but in search of glycogen in ascomycetes as such. (6-9): 582–591. TICHOMIROW A.W. 1908b. — Le glycogène des champignons asco- This late find of Hydnotrya confusa, with observations on its mycètes dans ses rapports avec la tréhalose. Bulletin des morphology and ecology, can possibly contribute to an awa- Sciences Pharmacologiques, 15: 189-196 + pl. V. reness that leads to further finds in the future. VRIES G.A. (DE) 2002. — Een vergelijkend onderzoek van de oliebol- truffel (Hydnotrya michaelis) met Hydnotrya cubispora en de voor Nederland nieuwe Hydnotrya confusa. Coolia, 45 (4): 201- Acknowlegements 204.

The authors thankfully acknowledge the support by A. Aku- lov, G. Hensel, B. Oertel, P. Reil and J.M. Trappe for fruitful discussions and for providing necessary literature, B. Spoo- ner for confirming the determination and H.O. Baral for re- viewing the manuscript. ef

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